Divergent responses of generalist and specialist pollinators to experimental drought: Outcomes for plant reproduction.

Drought is an increasingly important consequence of climate change. Drought often causes plants to alter patterns of resource allocation, which in turn can affect how plants interact with other species. How these altered interactions subsequently influence plant reproductive success remains incompletely understood and may depend on the degree of specialization exhibited by antagonists and mutualists. Specialist pollinators, for example, are dependent on floral resources from their obligate hosts and under drought conditions may thus indiscriminately visit these hosts (at least in certain circumstances). Generalist pollinators, in contrast, may only forage on host plants in good condition, given that they can forage on other plant species. We tested this hypothesis and its consequences for plant reproduction in squash (Cucurbita pepo) grown along an experimental moisture gradient ranging from dry (growth and flowering compromised) to wet conditions. Floral visitation increased with plant soil moisture for generalist honey bees but was independent of plant soil moisture for specialist squash bees. Pollen production increased with plant soil moisture, and fluorescent pigments placed on flowers revealed that pollinators primarily moved pollen from male flowers on well-watered plants to the stigmas of female flowers on well-watered plants. Seed set increased with increasing plant soil moisture but, notably, was higher in bee-pollinated plants compared to plants pollinated by hand with an even mix of pollen from plants grown at either end of the experimental moisture gradient. These results suggest that superior pollen rewards, perhaps combined with selective foraging by generalists, enhanced reproductive success in C. pepo when plant soil moisture was high and more generally illustrate that pollinator behavior may contribute to how drought conditions affect plant reproduction.

Genome-Wide Identification and Expression Analysis of Heat Shock Protein 70 (HSP70) Gene Family in Pumpkin (Cucurbita moschata) Rootstock under Drought Stress Suggested the Potential Role of these Chaperones in Stress Tolerance.

Heat shock protein 70s (HSP70s) are highly conserved proteins that are involved in stress responses. These chaperones play pivotal roles in protein folding, removing the extra amounts of oxidized proteins, preventing protein denaturation, and improving the antioxidant system activities. This conserved family has been characterized in several crops under drought stress conditions. However, there is no study on HSP70s in pumpkin (Cucurbita moschata). Therefore, we performed a comprehensive analysis of this gene family, including phylogenetic relationship, motif and gene structure analysis, gene duplication, collinearity, and promoter analysis. In this research, we found 21 HSP70s that were classified into five groups (from A to E). These genes were mostly localized in the cytoplasm, chloroplast, mitochondria, nucleus, and endoplasmic reticulum (ER). We could observe more similarity in closely linked subfamilies in terms of motifs, the number of introns/exons, and the corresponding cellular compartments. According to the collinearity analysis, gene duplication had occurred as a result of purifying selection. The results showed that the occurrence of gene duplication for all nine gene pairs was due to segmental duplication (SD). Synteny analysis revealed a closer relationship between pumpkin and cucumber than pumpkin and Arabidopsis. Promoter analysis showed the presence of various cis-regulatory elements in the up-stream region of the HSP70 genes, such as hormones and stress-responsive elements, indicating a potential role of this gene family in stress tolerance. We furtherly performed the gene expression analysis of the HSP70s in pumpkin under progressive drought stress. Pumpkin is widely used as a rootstock to improve stress tolerance, as well as fruit quality of cucumber scion. Since stress-responsive mobile molecules translocate through vascular tissue from roots to the whole plant body, we used the xylem of grafted materials to study the expression patterns of the HSP70 (potentially mobile) gene family. The results indicated that all CmoHSP70s had very low expression levels at 4 days after stress (DAS). However, the genes showed different expression patterns by progressing he drought period. For example, the expression of CmoHSP70-4 (in subgroup E) and CmoHSP70-14 (in subgroup C) sharply increased at 6 and 11 DAS, respectively. However, the expression of all genes belonging to subgroup A did not change significantly in response to drought stress. These findings indicated the diverse roles of this gene family under drought stress and provided valuable information for further investigation on the function of this gene family, especially under stressful conditions.

Integrating transcriptome and metabolome analyses of the response to cold stress in pumpkin (Cucurbita maxima).

Cucurbita maxima belong to the genus Cucurbita and are of nutritional and economic importance. Physiological activity, transcriptome, and metabolome analyses of leaf samples from the C. maxima inbreding line IL7 treated at 5 °C and 25 °C were performed. Cold stress resulted in a significant increase in the malondialdehyde content, relative electrical conductivity, soluble protein, sugar content, and catalase activity. A total of 5,553 differentially expressed genes were identified, of which 2,871 were up-regulated and 2,682 down-regulated. In addition, the transcription of differentially expressed genes in the plant hormone signal transduction pathway and transcription factor families of AP2/ERF, bHLH, WRKY, MYB, and HSF was activated. Moreover, 114 differentially expressed metabolites were identified by gas chromatography time-of-flight mass spectrometry, particularly through the analysis of carboxylic acids and derivatives, and organooxygen compounds. The demonstration of a series of potential metabolites and corresponding genes highlighted a comprehensive regulatory mechanism. These findings will provide novel insights into the molecular mechanisms associated with the response to cold stress in C. maxima.

Native and invasive ants affect floral visits of pollinating honey bees in pumpkin flowers (Cucurbita maxima).

Global pollinator decline is a major concern. Several factors-climate change, land-use change, the reduction of flowers, pesticide use, and invasive species-have been suggested as the reasons. Despite being a potential reason, the effect of ants on flowers received less attention. The consequences of ants being attracted to nectar sources in plants vary depending upon factors like the nectar source's position, ants' identity, and other mutualists interacting with the plants. We studied the interaction between flower-visiting ants and pollinators in Cucurbita maxima and compared the competition exerted by native and invasive ants on its pollinators to examine the hypothesis that the invasive ants exacerbate more interference competition to pollinators than the native ants. We assessed the pollinator's choice, visitation rate, and time spent/visit on the flowers. Regardless of species and nativity, ants negatively influenced all the pollinator visitation traits, such as visitation rate and duration spent on flowers. The invasive ants exerted a higher interference competition on the pollinators than the native ants did. Despite performing pollination in flowers with generalist pollination syndrome, ants can threaten plant-pollinator mutualism in specialist plants like monoecious plants. A better understanding of factors influencing pollination will help in implementing better management practices.

The role of alanine synthesis and nitrate-induced nitric oxide production during hypoxia stress in Cucurbita pepo nectaries.

Floral nectar is a sugary solution produced by nectaries to attract and reward pollinators. Nectar metabolites, such as sugars, are synthesized within the nectary during secretion from both pre-stored and direct phloem-derived precursors. In addition to sugars, nectars contain nitrogenous compounds such as amino acids; however, little is known about the role(s) of nitrogen (N) compounds in nectary function. In this study, we investigated N metabolism in Cucurbita pepo (squash) floral nectaries in order to understand how various N-containing compounds are produced and determine the role of N metabolism in nectar secretion. The expression and activity of key enzymes involved in primary N assimilation, including nitrate reductase (NR) and alanine aminotransferase (AlaAT), were induced during secretion in C. pepo nectaries. Alanine (Ala) accumulated to about 35% of total amino acids in nectaries and nectar during peak secretion; however, alteration of vascular nitrate supply had no impact on Ala accumulation during secretion, suggesting that nectar(y) amino acids are produced by precursors other than nitrate. In addition, nitric oxide (NO) is produced from nitrate and nitrite, at least partially by NR, in nectaries and nectar. Hypoxia-related processes are induced in nectaries during secretion, including lactic acid and ethanolic fermentation. Finally, treatments that alter nitrate supply affect levels of hypoxic metabolites, nectar volume and nectar sugar composition. The induction of N metabolism in C. pepo nectaries thus plays an important role in the synthesis and secretion of nectar sugar.

(3S,6E)-nerolidol-mediated rendezvous of Cyclocephala paraguayensis beetles in bottle gourd flowers.

Cyclocephalini beetles of the genus Cyclocephala (Coleoptera: Melolonthidae: Dynastinae) use flowers of some plants as food, shelter, and mating sites. However, little is known about floral scent chemistry involved in this interaction. Here we show that a sesquiterpene alcohol mediates attraction of Cyclocephala paraguayensis Arrow, on bottle gourd flowers, Lagenaria siceraria (Cucurbitaceae). Both males and females started to aggregate on the flowers at twilight; after that, mating began and remained for the entire night. GC-FID/EAD analysis of the L. siceraria floral scent collected in the field revealed that only the major constituent of the airborne volatiles elicited electroantennographic responses on male and female antennae of C. paraguayensis. This compound was identified as (3S,6E)-nerolidol, which was tested in two field trapping trials in Brazil. In the first bioassay, traps baited with nerolidol (mix of isomers) captured significantly more adult C. paraguayensis than control traps. In the second field trial, catches in traps baited with a mixture of isomers or enantiopure nerolidol were significantly higher than captures in control traps, but the treatments did not differ significantly. Analysis from the gut content of adult C. paraguayensis showed the presence of pollen, suggesting that they also use bottle gourd flowers for their nourishment. Taken together, these results suggest that (3S,6E)-nerolidol plays an essential role in the reproductive behavior of C. paraguayensis by eliciting aggregation, mating, and feeding.

Genome-wide characterization and expression analysis of the heat shock transcription factor family in pumpkin (Cucurbita moschata).

BACKGROUND: Crop quality and yield are affected by abiotic and biotic stresses, and heat shock transcription factors (Hsfs) are considered to play important roles in regulating plant tolerance under various stresses. To investigate the response of Cucurbita moschata to abiotic stress, we analyzed the genome of C. moschata. RESULTS: In this research, a total of 36 C. moschata Hsf (CmHsf) members were identified and classified into three subfamilies (I, II, and III) according to their amino acid sequence identity. The Hsfs of the same subfamily usually exhibit a similar gene structure (intron-exon distribution) and conserved domains (DNA-binding and other functional domains). Chromosome localization analysis showed that the 36 CmHsfs were unevenly distributed on 18 of the 21 chromosomes (except for Cm_Chr00, Cm_Chr08 and Cm_Chr20), among which 18 genes formed 9 duplicated gene pairs that have undergone segmental duplication events. The Ka/Ks ratio showed that the duplicated CmHsfs have mainly experienced strong purifying selection. High-level synteny was observed between C. moschata and other Cucurbitaceae species. CONCLUSIONS: The expression profile of CmHsfs in the roots, stems, cotyledons and true leaves revealed that the CmHsfs exhibit tissue specificity. The analysis of cis-acting elements and quantitative real-time polymerase chain reaction (qRT-PCR) revealed that some key CmHsfs were activated by cold stress, heat stress, hormones and salicylic acid. This study lays the foundation for revealing the role of CmHsfs in resistance to various stresses, which is of great significance for the selection of stress-tolerant C. moschata.

Citizen science improves our understanding of the impact of soil management on wild pollinator abundance in agroecosystems.

Native bees provide essential pollination services in both natural and managed ecosystems. However, declines in native bee species highlight the need for increased understanding of land management methods that can promote healthy, persistent populations and diverse communities. This can be challenging and costly using traditional scientific methods, but citizen science can overcome many limitations. In this study, we examined the distribution and abundance of an agriculturally important wild bee species, the squash bee (Eucera (Peponapis) pruinosa, Hymenoptera: Apidae). They are ground nesting, specialist bees that depend on cultivated varieties of Cucurbita (squash, pumpkins, gourds). The intimate relationship between squash bees and their host plants suggests that they are likely sensitive to farm management practices, particularly those that disturb the soil. In this study, citizen scientists across Michigan used a survey to submit field management and bee observation data. Survey results indicated that squash bees occupy a wide geographic range and are more abundant in farms with reduced soil disturbance. Citizen science provided an inexpensive and effective method for examining impacts of farm management practices on squash bees and could be a valuable tool for monitoring and conserving other native pollinators.

A light-induced decrease in the photochemical reflectance index (PRI) can be used to estimate the energy-dependent component of non-photochemical quenching under heat stress and soil drought in pea, wheat, and pumpkin.

The remote sensing of a plant's physiological state is a key problem of precision agriculture. The photochemical reflectance index (PRI), which is based on the intensities of the reflected light at 531 and 570 nm, is an important tool for the remote sensing of photosynthetic processes in plants. In particular, the PRI can be strongly connected with the non-photochemical quenching of chlorophyll fluorescence (NPQ) and the quantum yield of photosystem II (ФPSII); however, this connection is dependent on illumination, the intensity of stressor actions, the time scale of measurements, etc. The aim of the present work was to analyze the connection of PRI with the energy-dependent component of NPQ (NPQF) and ФPSII under heating and soil drought conditions. Pea, wheat, and pumpkin seedlings, which were grown under controlled conditions, were investigated. A PAM fluorometer Dual-PAM-100 and spectrometer S-100 were used for measurements of photosynthetic parameters and PRI, respectively. It was shown that heat stress increased the NPQF and the magnitude of light-induced changes in PRI (ΔPRI) and decreased ФPSII in pea seedlings. The decreased ФPSII and increased ΔPRI were observed in wheat after heating, but significant changes in NPQF were absent; the significant decrease in ФPSII was observed in pumpkin seedlings, while there were no significant changes in the other parameters. ΔPRI and NPQF after heating were significantly correlated. However, a significant correlation of the absolute values of PRI with photosynthetic parameters was absent. The soil drought increased NPQF and the magnitude of ΔPRI and decreased ФPSII in peas. ΔPRI was strongly correlated with photosynthetic parameters, but this correlation was absent for the absolute value of PRI. Thus, ΔPRI is strongly connected with the magnitude of NPQF and can be used as an estimator of this parameter.

Genomic Prediction of Pumpkin Hybrid Performance.

Genomic prediction has become an increasingly popular tool for hybrid performance evaluation in plant breeding mainly because that it can reduce cost and accelerate a breeding program. In this study, we propose a systematic procedure to predict hybrid performance using a genomic selection (GS) model that takes both additive and dominance marker effects into account. We first demonstrate the advantage of the additive-dominance effects model over the only additive effects model through a simulation study. Based on the additive-dominance model, we predict genomic estimated breeding values (GEBVs) for individual hybrid combinations and their parental lines. The GEBV-based specific combining ability (SCA) for each hybrid and general combining ability (GCA) for its parental lines are then derived to quantify the degree of midparent heterosis (MPH) or better-parent heterosis (BPH) of the hybrid. Finally, we estimate the variance components resulting from additive and dominance gene action effects and heritability using a genomic best linear unbiased predictor (g-BLUP) model. These estimates are used to justify the results of the genomic prediction study. A pumpkin ( spp.) data set is given to illustrate the provided procedure. The data set consists of 320 parental lines with 61,179 collected single nucleotide polymorphism (SNP) markers; 119, 120, and 120 phenotypic values of hybrids on three quantitative traits within maxima Duchesne; and 89, 111, and 90 phenotypic values of hybrids on the same three quantitative traits within Dechesne.

Impact of superparamagnetic iron oxide nanoparticles (SPIONs) and ionic iron on physiology of summer squash (Cucurbita pepo): A comparative study.

This study investigates the effect of SPIONs (superparamagnetic iron oxide nanoparticles, ∼12.5 nm in size) on summer squash plant (Cucurbita pepo) in the presence and absence of supplementary iron (Fe(II)-EDTA). The plants were grown in nutrient solution with different iron sources: (i) Fe(II)-EDTA, (ii) without Fe(II)-EDTA (iii) SPIONs only, and (iv) Fe(II)-EDTA with SPIONs. Plant growth and development were assessed after 20 days of soaking by measuring phenological parameters such as plant biomass, chlorophyll content, amount of carotenoids, and the catalase enzyme activity. Transmission electron microscopy, inductively coupled plasma atomic emission spectroscopy, X-ray diffraction, and vibrating sample magnetometer methods were used to detect uptake and translocation of SPIONs in plant tissues. Our results showed that SPIONs treatment (without Fe(II)-EDTA) caused growth retardation and decreased the plant biomass and chlorophyll content. Hence, they are not efficient sources to compensate for iron demand of squash plant. Electron microscopy observations, magnetization and elemental analyses revealed that SPIONs are taken-up by plant roots but not translocate to upper organs. In roots, SPIONs use a symplastic route for intercellular transfer. These findings suggest that as an iron source, SPIONs alone are not efficient for plant growth, but can contribute it together with Fe(II)-EDTA.

Spectroscopy can predict key leaf traits associated with source-sink balance and carbon-nitrogen status.

Approaches that enable high-throughput, non-destructive measurement of plant traits are essential for programs seeking to improve crop yields through physiological breeding. However, many key traits still require measurement using slow, labor-intensive, and destructive approaches. We investigated the potential to retrieve key traits associated with leaf source-sink balance and carbon-nitrogen status from leaf optical properties. Structural and biochemical traits and leaf reflectance (500-2400 nm) of eight crop species were measured and used to develop predictive 'spectra-trait' models using partial least squares regression. Independent validation data demonstrated that the models achieved very high predictive power for C, N, C:N ratio, leaf mass per area, water content, and protein content (R2>0.85), good predictive capability for starch, sucrose, glucose, and free amino acids (R2=0.58-0.80), and some predictive capability for nitrate (R2=0.51) and fructose (R2=0.44). Our spectra-trait models were developed to cover the trait space associated with food or biofuel crop plants and can therefore be applied in a broad range of phenotyping studies.

Uptake, transport, and effects of nano-copper exposure in zucchini (Cucurbita pepo).

Numerous studies on short term effects of copper-based nanomaterials on plants have been published, however investigations with plants grown in a complex soil medium are lacking. In this study Grey Zucchini (Cucurbita pepo) was grown in an environmental growth chamber using a 1:1 (v/v) potting mix native soil mixture amended with Kocide 3000, nCuO, bCuO, or Cu NPs. After 3 weeks Cu concentrations in the root, stem, and leaves of treated plants were significantly higher than control plants. This increase in Cu concentration did not adversely affect plant growth or chlorophyll production. The activity ascorbate peroxidase (APX) in the roots tissues of plants treated with Kocide 3000, nCuO, and bCuO decreased by at least 45%. Catalase (CAT) activity in root tissues of plants treated with 50 mg/kg of Cu NP decreased by 77%, while those treated at 200 mg/kg were reduced by 80%, compared to controls. The activity of APX and CAT in the leaves of all treated plants remained similar to control plants. Based on the endpoints used in this study, with the exception of a decrease in the accumulation of Zn and B in the roots, the exposure of zucchini to the tested copper compounds resulted in no negative effects.

First RNA-seq approach to study fruit set and parthenocarpy in zucchini (Cucurbita pepo L.).

BACKGROUND: Zucchini fruit set can be limited due to unfavourable environmental conditions in off-seasons crops that caused ineffective pollination/fertilization. Parthenocarpy, the natural or artificial fruit development without fertilization, has been recognized as an important trait to avoid this problem, and is related to auxin signalling. Nevertheless, differences found in transcriptome analysis during early fruit development of zucchini suggest that other complementary pathways could regulate fruit formation in parthenocarpic cultivars of this species. The development of next-generation sequencing technologies (NGS) as RNA-sequencing (RNA-seq) opens a new horizon for mapping and quantifying transcriptome to understand the molecular basis of pathways that could regulate parthenocarpy in this species. The aim of the current study was to analyze fruit transcriptome of two cultivars of zucchini, a non-parthenocarpic cultivar and a parthenocarpic cultivar, in an attempt to identify key genes involved in parthenocarpy. RESULTS: RNA-seq analysis of six libraries (unpollinated, pollinated and auxin treated fruit in a non-parthenocarpic and parthenocarpic cultivar) was performed mapping to a new version of C. pepo transcriptome, with a mean of 92% success rate of mapping. In the non-parthenocarpic cultivar, 6479 and 2186 genes were differentially expressed (DEGs) in pollinated fruit and auxin treated fruit, respectively. In the parthenocarpic cultivar, 10,497 in pollinated fruit and 5718 in auxin treated fruit. A comparison between transcriptome of the unpollinated fruit for each cultivar has been performed determining that 6120 genes were differentially expressed. Annotation analysis of these DEGs revealed that cell cycle, regulation of transcription, carbohydrate metabolism and coordination between auxin, ethylene and gibberellin were enriched biological processes during pollinated and parthenocarpic fruit set. CONCLUSION: This analysis revealed the important role of hormones during fruit set, establishing the activating role of auxins and gibberellins against the inhibitory role of ethylene and different candidate genes that could be useful as markers for parthenocarpic selection in the current breeding programs of zucchini.

An early ABA-induced stomatal closure, Na+ sequestration in leaf vein and K+ retention in mesophyll confer salt tissue tolerance in Cucurbita species.

Tissue tolerance to salinity stress is a complex physiological trait composed of multiple 'sub-traits' such as Na+ compartmentalization, K+ retention, and osmotic tolerance. Previous studies have shown that some Cucurbita species employ tissue tolerance to combat salinity and we aimed to identify the physiological and molecular mechanisms involved. Five C. maxima (salt-tolerant) and five C. moschata (salt-sensitive) genotypes were comprehensively assessed for their salt tolerance mechanisms and the results showed that tissue-specific transport characteristics enabled the more tolerant lines to deal with the salt load. This mechanism was associated with the ability of the tolerant species to accumulate more Na+ in the leaf vein and to retain more K+ in the leaf mesophyll. In addition, C. maxima more efficiently retained K+ in the roots when exposed to transient NaCl stress and it was also able to store more Na+ in the xylem parenchyma and cortex in the leaf vein. Compared with C. moschata, C. maxima was also able to rapidly close stomata at early stages of salt stress, thus avoiding water loss; this difference was attributed to higher accumulation of ABA in the leaf. Transcriptome and qRT-PCR analyses revealed critical roles of high-affinity potassium (HKT1) and intracellular Na+/H+ (NHX4/6) transporters as components of the mechanism enabling Na+ exclusion from the leaf mesophyll and Na+ sequestration in the leaf vein. Also essential was a higher expression of NCED3s (encoding 9-cis-epoxycarotenoid dioxygenase, a key rate-limiting enzyme in ABA biosynthesis), which resulted in greater ABA accumulation in the mesophyll and earlier stomata closure in C. maxima.

Physiological response of Cucurbita pepo var. pepo mycorrhized by Sonoran desert native arbuscular fungi to drought and salinity stresses

ABSTRACT Plants response to symbiosis with arbuscular mycorrhizal fungi (AMF) under water stress is important to agriculture. Under abiotic stress conditions native fungi are more effective than exotics in improving plant growth and water status. Mycorrhization efficiency is related to soil fungi development and energy cost-benefit ratio. In this study, we assessed the effect on growth, water status and energy metabolism of Cucurbita pepo var. pepo when inoculated with native AMF from the Sonoran desert Mexico (mixed isolate and field consortium), and compared with an exotic species from a temperate region, under drought, low and high salinity conditions. Dry weights, leaf water content, water and osmotic potentials, construction costs, photochemistry and mycorrhization features were quantified. Under drought and low salinity conditions, the mixed isolate increased plant growth and leaf water content. Leaf water potential was increased only by the field consortium under drought conditions (0.5-0.9 MPa). Under high salinity, the field consortium increased aerial dry weight (more than 1 g) and osmotic potential (0.54 MPa), as compared to non-mycorrhized controls. Plants inoculated with native AMF, which supposedly diminish the effects of stress, exhibited low construction costs, increased photochemical capacity, and grew larger external mycelia in comparison to the exotic inoculum.

Pumpkin as an Alternate Host Plant for Laboratory Colonies of Grape Mealybug.

The grape mealybug, Pseudococcus maritimus (Ehrhorn; Hemiptera: Pseudococcidae), is the primary vector of Grapevine Leafroll associated Viruses (GLRaVs) in Washington State vineyards. Rearing laboratory colonies of grape mealybug has proven difficult. Several host plants were tested to determine their suitability for use as an alternate host plants for laboratory colonies of grape mealybug. Of the plants tested, colonies of grape mealybug were successful on pumpkin (Cucurbita pepo cv 'Connecticut field') leaves and vines. Mealybugs were able to develop from the crawler stage, through developmental instars, and adults were able to reproduce viable offspring. To date this is the only successful study to raise grape mealybugs on a cucurbit. Pumpkin appears to be a viable alternate host plant for laboratory colonies of the grape mealybug.

Physiological response of Cucurbita pepo var. pepo mycorrhized by Sonoran desert native arbuscular fungi to drought and salinity stresses.

Plants response to symbiosis with arbuscular mycorrhizal fungi (AMF) under water stress is important to agriculture. Under abiotic stress conditions native fungi are more effective than exotics in improving plant growth and water status. Mycorrhization efficiency is related to soil fungi development and energy cost-benefit ratio. In this study, we assessed the effect on growth, water status and energy metabolism of Cucurbita pepo var. pepo when inoculated with native AMF from the Sonoran desert Mexico (mixed isolate and field consortium), and compared with an exotic species from a temperate region, under drought, low and high salinity conditions. Dry weights, leaf water content, water and osmotic potentials, construction costs, photochemistry and mycorrhization features were quantified. Under drought and low salinity conditions, the mixed isolate increased plant growth and leaf water content. Leaf water potential was increased only by the field consortium under drought conditions (0.5-0.9MPa). Under high salinity, the field consortium increased aerial dry weight (more than 1g) and osmotic potential (0.54MPa), as compared to non-mycorrhized controls. Plants inoculated with native AMF, which supposedly diminish the effects of stress, exhibited low construction costs, increased photochemical capacity, and grew larger external mycelia in comparison to the exotic inoculum.

Root respiratory burst oxidase homologue-dependent H2O2 production confers salt tolerance on a grafted cucumber by controlling Na+ exclusion and stomatal closure.

Plant salt tolerance can be improved by grafting onto salt-tolerant rootstocks. However, the underlying signaling mechanisms behind this phenomenon remain largely unknown. To address this issue, we used a range of physiological and molecular techniques to study responses of self-grafted and pumpkin-grafted cucumber plants exposed to 75 mM NaCl stress. Pumpkin grafting significantly increased the salt tolerance of cucumber plants, as revealed by higher plant dry weight, chlorophyll content and photochemical efficiency (Fv/Fm), and lower leaf Na+ content. Salinity stress resulted in a sharp increase in H2O2 production, reaching a peak 3 h after salt treatment in the pumpkin-grafted cucumber. This enhancement was accompanied by elevated relative expression of respiratory burst oxidase homologue (RBOH) genes RbohD and RbohF and a higher NADPH oxidase activity. However, this increase was much delayed in the self-grafted plants, and the difference between the two grafting combinations disappeared after 24 h. The decreased leaf Na+ content of pumpkin-grafted plants was achieved by higher Na+ exclusion in roots, which was driven by the Na+/H+ antiporter energized by the plasma membrane H+-ATPase, as evidenced by the higher plasma membrane H+-ATPase activity and higher transcript levels for PMA and SOS1. In addition, early stomatal closure was also observed in the pumpkin-grafted cucumber plants, reducing water loss and maintaining the plant's hydration status. When pumpkin-grafted plants were pretreated with an NADPH oxidase inhibitor, diphenylene iodonium (DPI), the H2O2 level decreased significantly, to the level found in self-grafted plants, resulting in the loss of the salt tolerance. Inhibition of the NADPH oxidase-mediated H2O2 signaling in the root also abolished a rapid stomatal closure in the pumpkin-grafted plants. We concluded that the pumpkin-grafted cucumber plants increase their salt tolerance via a mechanism involving the root-sourced respiratory burst oxidase homologue-dependent H2O2 production, which enhances Na+ exclusion from the root and promotes an early stomatal closure.

Translocation of heavy metals from soils into floral organs and rewards of Cucurbita pepo: Implications for plant reproductive fitness.

Metals and metalloids in soil could be transferred into reproductive organs and floral rewards of hyperaccumulator plants and influence their reproductive success, yet little is known whether non-hyperaccumulator plants can translocate heavy metals from soil into their floral organs and rewards (i.e., nectar and pollen) and, if so, whether plant reproduction will be affected. In our studies, summer squash (Cucurbita pepo L. cv. Golden Apple) was exposed to heavy-metal treatments during bud stage to investigate the translocation of soil-supplemented zinc, copper, nickel and lead into its floral organs (pistil, anther and nectary) and rewards (nectar and pollen) as well as floral metal accumulation effects on its reproduction. The results showed that metals taken up by squash did translocate into its floral organs and rewards, although metal accumulation varied depending on different metal types and concentrations as well as floral organ/reward types. Mean foraging time of honey bees to each male and female flower of squash grown in metal-supplemented soils was shorter relative to that of plants grown in control soils, although the visitation rate of honeybees to both male and female flowers was not affected by metal treatments. Pollen viability, pollen removal and deposition as well as mean mass per seed produced by metal-treated squash that received pollen from plants grown in control soils decreased with elevated soil-supplemented metal concentrations. The fact that squash could translocate soil-supplemented heavy metals into floral organs and rewards indicated possible reproductive consequences caused either directly (i.e., decreasing pollen viability or seed mass) or indirectly (i.e., affecting pollinators' visitation behavior to flowers) to plant fitness.